The present invention relates to seeding crystals for the preparation of peptides or proteins such as zinc insulin products.
The xe2x80x9cLentexe2x80x9d family of zinc insulin products are insulin zinc suspensions of the type originally developed in the 1950""s with the aim of producing insulin preparations that would be able to cover diabetics"" insulin requirement with a single daily injection (see erg. Jens Brange, Galenics of Insulin, 1987). Various Lente insulin products having different action profiles are available in the form of different combinations of amorphous and/or crystalline insulin particles from Novo Nordisk A/S, Denmark These include SEMILENTE, a suspension of amorphous insulin particles, ULTRALENTE, a suspension of crystalline insulin particles, and LENTE, which is a mixture of 30% amorphous and 70% crystalline insulin particles.
For several decades, seeding crystals for preparation of the xe2x80x9cLentexe2x80x9d zinc insulin products have been prepared by the same basic freeze-drying method that was developed and patented in the early 1950""s. This method, which is described in GB patent specification No. 766,994, involves the addition of freeze-dried amorphous insulin, typically beef insulin, to an insulin-containing crystallization medium to result in the formation of a suspension of insulin microcrystals of a size of about 2-7 xcexcm. This suspension, which is eventually used for the preparation of the final crystalline zinc insulin product, is filled into small vials (e.g. 10 ml), frozen in an alcohol/carbon dioxide mixture and stored frozen at a temperature at or below xe2x88x9218xc2x0 C.
Although still in use, this method has a number of disadvantages:
1. It is based on the use of beef insulin, since it has until now not been possible to produce acceptable microcrystals of pure human insulin. As a result of the use of beef insulin nuclei for the formation of the microcrystals, the end product contains a small amount of beef insulin, which is undesirable.
2. The freeze-drying method requires a lyophilizer and subsequent shipping and storage at a temperature of no more than xe2x88x9218xc2x0 C. This is expensive and requires a great deal of space.
3. The method of preparation is extremely difficult to perform in a sufficiently aseptic manner.
It would therefore be advantageous to be able to produce insulin seeding crystals using a method which does not suffer from the disadvantages of the known methods. It has now surprisingly been found that it is possible, using a relatively simple and inexpensive process, to produce insulin seeding crystals which are free of beef insulin, which can be stored at room temperature and which result in insulin preparations having advantageous properties in terms o,f e.g., crystal particle size and uniformity. Furthermore, it is also contemplated that this process will be applicable to the production of seeding crystals for other peptides and proteins, in particular peptides or proteins used as pharmaceuticals.
It is thus an object of the present invention to provide a novel method for the production of peptide or protein seeding crystals. More particularly, it is an object of the invention to provide a method for the production of insulin seeding crystals which does not require the use of beef insulin, which makes possible storage and transport without the need for expensive freeze-drying and storage at sub-zero temperatures, and which can be performed in a closed system so as to more readily allow the use of aseptic production methods.
Another object of the invention is to provide a method for the preparation of insulin seeding crystals for the production of crystalline zinc insulin suspensions having a narrow particle size distribution.
In its broadest aspect, the present invention thus relates to a method for producing seeding microcrystals for the production of a peptide or protein, comprising providing an unseeded suspension of a peptide or protein and homogenizing said suspension under pressure to result in peptide or protein microcrystals suitable for use as seeding microcrystals.
In a particular embodiment, the invention relates to a method for producing seeding microcrystals for the production of human insulin, said microcrystals being free of non-human pancreatic insulin, comprising providing an unseeded suspension of human insulin, said suspension being free of non-human pancreatic insulin, and homogenizing said insulin suspension under pressure to result in human insulin microcrystals suitable for use as seeding microcrystals for the production of zinc insulin products.
Another aspect of the invention relates to a method for the production of a peptide or protein product, comprising providing an unseeded suspension of a peptide or protein and seeding said suspension with microcrystals produced by the method indicated above.
In a particular embodiment of this aspect of the invention, the peptide or protein product to be produced is a zinc insulin product, and the unseeded suspension is a suspension of human insulin.
A further aspect of the invention relates to human insulin microcrystals suitable for use as seeding microcrystals for the production of zinc insulin products, said microcrystals being free of non-human pancreatic insulin.
A still further aspect of the invention relates to human zinc insulin product free of non-human pancreatic insulin.